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Chemical Composition and in Vitro Antibacterial Activity of Essential Oil

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Chemical Composition and in Vitro Antibacterial Activity of Essential Oil American Journal of Essential Oils and Natural Products 2017; 5(3): 01-06 ISSN: 2321-9114 AJEONP 2017; 5(3): 01-06 Chemical composition and in vitro antibacterial activity © 2017 AkiNik Publications Received: 02-05-2017 of essential oil from Eupatorium africanum Oliv. & Accepted: 04-06-2017 Hiern Philippe Babady-Bila Bila Nutraceuticals, 650 Portland Street Market, Philippe Babady-Bila, Dorothée Tshilanda Dinangayi, Damien Sha- Dartmouth, NS B2W 6P3, Tshibey Tshibangu, Emmanuel Lengbiye, Jean-Paul Ngbolua and Pius Canada Mpiana Tshimankinda Dorothée Tshilanda Dinangayi Department of Chemistry, Abstract University of Kinshasa, Kin. XI A hydrodistilled oil from the aerial part of Eupatorium africanum Oliv. & Hiern collected from D.R. Kinshasa, D.R. Congo Congo was analyzed by GC and GC/MS. Thirty three compounds representing 97.34% of the oil were Damien Sha-Tshibey Tshibangu identified. The major components were -eudesmol (49.1%), 10-epi--eudesmol (11.30%), -humulene Department of Chemistry, (8.9%), hedycaryol (8.9%), and elemol (5.2%). The antibacterial activity of the essential oil and its University of Kinshasa, Kin. XI dominant constituent was evaluated against four bacteria strains (Staphyloccocus aureus ATCC 25952, Kinshasa, D.R. Congo Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 9027, Proteus vulgaris ATCC 4635) using the micro-dilution method. The results indicate that the two samples are active against all tested Emmanuel Lengbiye microorganisms. The oil has a moderate activity against Staphylococcus aureus and Pseudomonas Department of Biology, aeruginosa with a minimum inhibitory concentration (MIC) value of 125 µg/mL. The isolated major University of Kinshasa, Kin. XI constituent (-eudesmol) exhibits the highest activity against Staphylococcus aureus and Pseudomonas Kinshasa, D.R. Congo aeruginosa with a MIC value of 62.5 µg/mL. Both samples exhibited a MIC value of 250 μg/ml against Escherichia coli. and Proteus vulgaris. Jean-Paul Ngbolua Department of Biology, Keywords: Eupatorium africanum Oliv. & Hiern, Asteraceae, essential oil, chemical composition, University of Kinshasa, Kin. XI Kinshasa, D.R. Congo biological activity. Pius Mpiana Tshimankinda 1. Introduction Department of Chemistry, Eupatorium africanum Oliv. & Hiern. [syn. Stomatanthes africanus (Oliv. & Hiern) R.M. University of Kinshasa, Kin. XI King & H. Rob] belongs to the genus Stomatanthes, Asteraceae family, Eupatorieae tribe. This Kinshasa, D.R. Congo genus comprises 17 species distributed in South America (13 species) and in Africa (4 species). But according to Grossi, the genus Stomatanthes would be formed by only three of the four African species (S. africanus, S. meyeri and S. helenae). Stomatanthes zambiensis, the fourth African species, is excluded from Stomatanthes genus. It would be related with Eupatorium sensu stricto [1]. This plant is a perennial herb (sub-shrub 20–120 cm high) of hilly grassland throughout the [2] region from Guinea to West Cameroon and widespread in tropical Africa . It is used as herbal medicine against headache, diarrhea, dysentery and naso-pharyngeal affections [2, 3]. In Congo, a leaf decoction is made as a mouthwash for aphthera, and a tisane of the boiled root is used as a cough medicine and an anti-diarrhetic [4]. Phytochemical investigations and chemical composition of Eupatorium species essential oils (including many species now reclassified into other closely related genera) have been carried out by several researchers. These studies revealed the presence of a variety of monoterpernes, sesquiterpenes, sesquiterpene lactones, flavonoids, phenolic and acetylenic compounds, triterpenes, and alkaloids with cytotoxic, antitumoral, antimicrobial, antioxidant and anti- [5-6] inflammatory activities has been reported . A review of antimicrobial activity of the genus Eupatorium L. (Asteraceae) has been recently published and Eupatorium africanum is not mentioned in this paper [7]. The following terpenoids were reported as major constituents of Correspondence: Eupatorium species essential oils: α-pinene (50.98 %), -pinene (10.5%) sabinene (46.7%), Philippe Babady-Bila limonene (9.63-23.3%), camphene (8.9%), (+)-camphor (15.46%), 2,5-dimethoxy-p-cymene Bila Nutraceuticals, 650 Portland Street Market, or thymohydroquinone dimethyl ether (20.8-60.7%), thymol, p-cymene (23.7%), thymol Dartmouth, NS B2W 6P3, methyl ether (36.3%), aristolone+laevigatin (23.6%), globulol (16.2-25.1%), caryophyllene Canada oxide (17.4–30.1%), germacrene D (8.6–21.6%), spathulenol (14.2%), (-)-spathulenol ~ 1 ~ American Journal of Essential Oils and Natural Products (0.48-25.16 %), α-zingiberene (57.5%), α-gurjunene (19.5%), operated at an ionization energy of 70 eV with a mass scan (E)-β-bisabolene (9.7%), α-selinene (9.0%), β-caryophyllene range of 40-400 amu. (1.11-41.7%), ocimene (4.78 %), δ-elemene (10.57%), α- humulene (0.25-14.6%), patchoulene (9.24%), viridiflorol 2.5 Identification of oil constituents (9.16%), γ-elemene (5.92%), (+)-δ-cadinene (5.83%), (-)-δ- The identification of constituents was based on a comparison cadinol (2.67%), α-santalene (2.53%), β-cubebene (1.64%), of their retention indices and mass spectra with spectral data (+)-nerolidol (1.63%), selina-4(15),7(11)-dien-8-one (36.6%), from several sources, by searching in the GC-MS library, and -muurolene (10.4-19%), myrcene (15.7%), valencene by matching their fragmentation patterns in mass spectra with (10.5%), cyperone (16.9%), and pregeijerene (14.3%), those of published [24-28]. The percentage of the individual phellandrene (3.85%), α-bisabolol (9.53%), bornyl acetate constituent was obtained from FID area percentages without (8.98-15.6%), -bisabolene (6.16%), amorph-4-en-7-ol the use of correction factors. Retention indices were (9.6%), 3-acetoxyamorpha-4,7(11)- dien-8-one (7.8%) and calculated relative to C8-C24 n-alkanes, and compared with amorph-4,7(11)-dien-8-one (5.7%), neryl isobutyrate (17.6%). values reported in the literature [24-26]. The major non-terpenoid compounds reported were methyl chavicol (42.2%) and 1-naphthalenol (17.50%). 2.6 Antibacterial Activity Amorph-4-en-7-ol (9.6%), 3-acetoxyamorpha - 4,7(11) - dien- 2.6.1 Microorganisms 8-one (7.8%) and amorph-4,7(11)-dien-8-one (5.7%) were Four strains of microorganisms from the American Type identified in Eupatorium adenophorum oil. Amorphene Culture Collection (ATCC, Rockville MD, USA) were used derivatives (19.8–41.4%) may be considered as characteristic in the present study: Staphylococcus aureus (S. aureus ATCC constituents of E. adenophorum [5, 8-23]. 25952), Escherichia coli (E. coli ATCC 27195), The present investigation reports the results of GC and GC- Pseudomonas aeruginosa (P. aeruginosa ATCC 9027), and MS analyses of the essential oil from the leaves of Proteus vulgaris (P. vulgaris ATCC 4635) strains. Eupatorium africanum Oliv. & Hiern growing wild in D.R. Congo, and its antibacterial activity evaluation. To the best of 2.6.2 Determination of Minimum inhibitory concentration our knowledge, this is the first report on the chemical (MIC) composition and the biological activity of the The antibacterial activity of Eupatorium africanum essential Eupatorium africanum essential oil. oil and its major constituent was assessed against selected bacteria strains by the micro-well dilution method and the 2. Materials and methods minimum inhibitory concentration (MIC) values, which 2.1 Plant material represent the lowest sample concentrations that completely The Eupatorium africanum leaves were collected in the inhibit the growth of microorganisms were obtained using this Bombo-Lumene Game reserve, Kinshasa Province, D.R. method [29]. Congo, and authenticated by a voucher specimen (H. The 10 mg samples (essential oil and isolated major BREYNE 2785) kept at the INERA herbarium, Department of constituent) were each dissolved in DMSO (250 μL) and Biology, University of Kinshasa, D.R. Congo. diluted with Mueller–Hinton Broth (MHB) in order to reach The collected fresh leaves were dried at room temperature in a concentrations of 2000 μg/mL and a 5 ml solution (final well-ventilated room and preserved in tightly closed bumper volume, and 5% DMSO final concentration). These solution bags at room temperature in the absence of light. were used as stock solutions. The inocula of microorganisms were prepared from 24h old 2.2 Isolation of essential oil MHB cultures. The microbial suspensions were prepared by Dried plant material (300 g) was subjected to hydro- adding five colonies of each of the test bacteria to 2 mL of distillation for 4 hours using a Clevenger type apparatus to with sterile physiological solution (0.9% NaCl) and adjusted produce an essential oil at a yield of 0.4%. The oil was dried with this sterile physiological solution to match that of a 0.5 over anhydrous sodium sulfate and stored in sealed vials at McFarland standard solution (108 cells/ml). They were then low temperature (4 C). diluted (1/100) to achieve 106 CFU/mL. The assay was carried out using sterile clear polystyrene 96- 2.3 Gas Chromatographic (GC) Analysis well microtiter plates (round bottom). The wells in the GC analysis was performed using a Hewlett-Packard 5880A columns 2 to 8 and those in columns 11 and 12 were filled gas chromatograph equipped with a flame ionization detector with 100 µL MHB (Mueller Hinton Broth). Briefly 200 µL of (FID) and data-handling system. The GC was fitted with a stock solution of each Eupatorium sample were added to the DB-5 fused silica capillary column (30m × 0.25 mm, film wells in column 1 (A1 to H1), and two-fold serial dilutions thickness 0.25 μm). The oven temperature was programmed were made from column 1 to column 8. Then 5 µL of the as follows: 40 °C for 5 min, from 40 to 200 °C at a rate of 4 inoculum were dispensed to all the wells except those in °C/ min, and 10 oC/min. ramp to 300 oC.
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